Detection of carcinoembryonic antigen using functional magnetic and fluorescent nanoparticles in magnetic separators

We combined a sandwich immunoassay, anti-CEA/CEA/anti-CEA, with functional magnetic (~80 nm) and fluorescent (~180 nm) nanoparticles in magnetic separators to demonstrate a detection method for carcinoembryonic antigen (CEA). Determination of CEA in serum can be used in clinical diagnosis and monitoring of tumor-related diseases. The CEA concentrations in samples were deduced and determined based on the reference plot using the measured fluorescent intensity of sandwich nanoparticles from the sample. The linear range of CEA detection was from 18 ng/mL to 1.8 pg/mL. The detection limit of CEA was 1.8 pg/mL. In comparison with most other detection methods, this method had advantages of lower detection limit and wider linear range. The recovery was higher than 94%. The CEA concentrations of two serum samples were determined to be 9.0 and 55 ng/mL, which differed by 6.7% (9.6 ng/mL) and 9.1% (50 ng/mL) from the measurements of enzyme-linked immunosorbent assay (ELISA), respectively. The analysis time can be reduced to one third of ELISA. This method has good potential for other biomarker detections and biochemical applications.     

A Novel and Highly Sensitive Resonance Scattering Spectral Assay for Horseradish Peroxidase Using Cationic Surfactant

A novel and ultrasensitive resonance scattering (RS) spectral assay was proposed for detection of horseradish peroxidase (HRP) activity. It was based on that the HRP strongly catalyze H₂O₂ oxidation of excess I⁻ to form , the resulting combined with four cationic surfactant (CS), including tetradecyl pyridinium bromide (TPB), cetyltrimethyl ammonium bromide (CTMA), cetylpyridinium chloride (CPCM) and tetradecyl dimethylbenzyl ammonium chloride (TDMBA) to produce association particles (TPB-I₃) _n , (CTMA-I₃) _m , (CPCM-I₃)_l and (TDMBA-I₃) _k , which exhibit a strongest resonance scattering peak at 478, 423, 538 and 491 nm, respectively. For the four systems of TPB, CPCM, CTMBA and TDMBA, the HRP activity determined was in the linear range of 0.004–5.6, 0.04–3.2, 0.04–8.0, 0.08–8.0 ng/mL, with a detection limit of 0.0034, 0.040, 0.033, 0.016 ng/mL, respectively. The TPB resonance scattering spectral assay was best and has been applied to the analysis of HRP in real samples, with satisfactory results.     

The Application of CdTe/CdS in the Detection of Carcinoembryonic Antigen by Fluorescence Polarization Immunoassay

A novel and portable strategy based on fluorescence polarization immunoassay (FPIA) using quantum dots (QDs) was described in this study for simple, rapid, and sensitive detection of carcinoembryonic antigen (CEA). Under optimal conditions, the sensor has a wide dynamic range (from 0.5?ng/mL to 200?ng/mL) and a good correlation. The limit of detection (LOD) is 0.21?ng/mL (S/N?=?3). The sensor has been applied for detection of carcinoembryonic antigen in 10 human serum samples with the range of recovery from 92.1?% to 103.6?%. Furthermore, bioconjugation of the core–shell QDs with streptavidin (SA) has been successfully applied in immunofluorescent imaging of the human hepatocellular carcinoma (HEPG2) cell line. The experimental results demonstrated the successful application of QDs-based fluorescence polarization immunoassay for detection of target proteins of biomedical importance. This strategy shows great promise for clinical diagnoses and basic discovery with high sensitivity, good specificity, simple procedures and short analysis time.     

Green sonochemical synthesis and fabrication of cubic MnFe2O4 electrocatalyst decorated carbon nitride nanohybrid for neurotransmitter detection in serum samples

The binary nanomaterials and graphitic carbon based hybrid has been developed as an important porous nanomaterial for fabricating electrode with applications in non-enzymatic (bio) sensors. We report a fast synthesis of bimetal oxide particles of nano-sized manganese ferrite (MnFe₂O₄) decorated on graphitic carbon nitride (GCN) via a high-intensity ultrasonic irradiation method for C (30 kHz and 70 W/cm²). The nanocomposites were analyzed by powder X-ray diffraction, XPS, EDS, TEM to ascertain the effects of synthesis parameters on structure, and morphology. The MnFe₂O₄/GCN modified electrode demonstrated superior electrocatalytic activity toward the neurotransmitter (5-hydroxytryptamine) detection with a high peak intensity at +0.21 V. The appealing application of the MnFe₂O₄/GCN/GCE as neurotransmitter sensors is presented and a possible sensing mechanism is analyzed. The constructed electrochemical sensor for the detection of 5-hydroxytryptamine (STN) showed a wide working range (0.1–522.6 μM), high sensitivity (19.377 μA μM⁻¹ cm⁻²), and nano-molar detection limit (3.1 nM). Moreover, it is worth noting that the MnFe₂O₄/GCN not only enhanced activity and also promoted the electron transfer rate towards STN detection. The proposed sensor was analyzed for its real-time applications to the detection of STN in rat brain serum, and human blood serum in good satisfactory results was obtained. The results showed promising reproducibility, repeatability, and high stability for neurotransmitter detection in biological samples.     

增强型酶催化发光免疫分析法高灵敏测定血清中心肌肌钙蛋白Ⅰ

基于辣根过氧化物酶（HRP）催化 H₂O₂-Luminol系统,对一种新型酚类衍生物 4-（1,2,4-三氮唑-1-基）苯酚对化学发光的增强作用进行了研究。用HRP标记急性心肌梗死标志物心肌肌钙蛋白Ⅰ（cTnⅠ）单克隆抗体,通过cTnⅠ的双抗体夹心免疫反应,建立了简单、灵敏和快速检测人血清中的cTnⅠ含量的增强型酶发光免疫分析方法。实验结果表明,cTnⅠ在1.2 ~ 24 ng/mL浓度范围内与增强型发光强度具有良好的线性关系（R=0.99）,变异系数（n= 8）为 4.7%。最后,使用该方法对人血清样品中的cTnⅠ含量进行测定,测试结果具有较好的稳定性和精度,能够满足临床检测要求。     

An orientation analysis method for protein immobilized on quantum dot particles

The evaluation of orientation of biomolecules immobilized on nanodevices is crucial for the development of high performance devices. Such analysis requires ultra high sensitivity so as to be able to detect less than one molecular layer on a device. Time-of-flight secondary ion mass spectrometry (TOF-SIMS) has sufficient sensitivity to evaluate the uppermost surface structure of a single molecular layer. The objective of this study is to develop an orientation analysis method for proteins immobilized on nanomaterials such as quantum dot particles, and to evaluate the orientation of streptavidin immobilized on quantum dot particles by means of TOF-SIMS. In order to detect fragment ions specific to the protein surface, a monoatomic primary ion source (Ga⁺) and a cluster ion source (Au₃⁺) were employed. Streptavidin-immobilized quantum dot particles were immobilized on aminosilanized ITO glass plates at amino groups by covalent bonding. The reference samples streptavidin directly immobilized on ITO plates were also prepared. All samples were dried with a freeze dryer before TOF-SIMS measurement. The positive secondary ion spectra of each sample were obtained using TOF-SIMS with Ga⁺ and Au₃⁺, respectively, and then they were compared so as to characterize each sample and detect the surface structure of the streptavidin immobilized with the biotin-immobilized quantum dots. The chemical structures of the upper surface of the streptavidin molecules immobilized on the quantum dot particles were evaluated with TOF-SIMS spectra analysis. The indicated surface side of the streptavidin molecules immobilized on the quantum dots includes the biotin binding site.     

An electrochemical sensing platform based on a new Cd(II)-containing ionic liquid for the determination of trichloroacetic acid and bromate

The present work reports on the synthesis, characterization and performance of a new metal-containing ionic liquid [(C₃H₇)₂-bim]₂[CdCl₄] (bim = benzimidazole) as an electrocatalyst for trichloroacetic acid (TCA) and bromate reduction. The structure of Cd(II)-containing ionic liquid (Cd-IL) was characterized by X-ray crystallography, IR spectroscopy, and elemental analysis. The molecular structure contains two independent cations of 1,3-dipropyl-benzimidazolium and one anion of CdCl₄²⁻. The cadmium atom has a tetrahedral geometry by coordinating to four chlorine atoms. The melting point of Cd-IL is 73 °C. Electrochemical properties of the Cd-IL have been investigated by preparing bulk-modified carbon paste electrode, and Cd-IL is used as a binder and an electrocatalyst. This modified electrode has good electrocatalytic activity toward reduction of TCA and bromate. The detection limit and the sensitivity are 0.01 μM and 102.72 μA μM⁻¹ for trichloroacetic acid detection and 0.003 μM and 496.15 μA μM⁻¹ for bromate detection. This work demonstrates that the Cd-IL may become a new kind of functional material in constructing chemicals and biosensors.     

ZrO<Subscript>2</Subscript>/DNA-derivated polyion hybrid complex membrane for the determination of hydrogen peroxide in milk

An efficient biosensing substrate based on ZrO₂/DNA-derivated polyion complex (PIC) membrane has been developed for the determination of hydrogen peroxide (H₂O₂) in this study. To fabricate such a PIC membrane, ZrO₂ nanoparticles were initially electrodeposited on the bare gold electrode (ZrO₂/Au), and deoxyribonucleic acid (DNA)-doped hemoglobin mixture was then assembled onto the ZrO₂/Au surface. The double-strand DNA provided a biocompatible microenvironment for the immobilization of biomolecules, greatly amplified the surface coverage of biomolecules on the electrode surface, and improved the sensitivity of the biosensor. The fabricated procedure of the proposed biosensor was characterized by cyclic voltammetry, electrochemical impedance spectroscopy, and atomic force microscopy. The performance and factors influencing the performance of the biosensor were also evaluated. Under optimal conditions, the developed biosensor exhibited a well-defined electrochemical behavior toward the reduction of H₂O₂ ranging from 1.1 μM to 2.3 mM with a detection limit of 0.5 μM (S/N = 3). The biosensor was applied to the determination of H₂O₂ in milk with satisfactory results. It is important to note that the PIC membrane provided an alternative substrate for the immobilization of other proteins.     

Nano TiO2 film electrode for electrocatalytic reduction of furfural in ionic liquids

Nanoporous TiO₂ having enhanced surface area was synthesized by sol–gel method. An “environmental friendly” method for production of furfuryl alcohol was presented by electrocatalytic reduction of furfural to furfuryl alcohol in ionic liquid medium at the surface of nanoporous TiO₂ film electrode. The heterogeneous catalytic redox behaviour of a nanoporous TiO₂ film electrode surface was investigated by cyclic voltammetry (CV). It was found that the catalytic reduction of furfural by Ti(IV)/Ti(III) redox system on the nanoporous TiO₂ film surface. The electrode reaction mechanism is called catalytic (EC′) mechanism, current density can reach 38 mA/cm² and yielding an overall conversion efficiency of 61.7%.     

Electrical studies on ionic liquid-based gel polymer electrolyte for its application in EDLCs

Ionic liquid-based gel polymer electrolyte (GPE) has been synthesized using standard solution cast technique. Different weight percent of ionic liquid, 1-Butyl-3-methylimidazolium chloride (BMIMCl) and liquid electrolyte, ethylene carbonate (EC)–propylene carbonate (PC)–tetra ethyl ammonium tetra fluoro borate (TEABF₄) was incorporated in polymer, poly(vinylidene fluoride-co-hexafluoro propylene (PVdF-HFP) to obtain mechanically stable gel polymer electrolyte film (GPE) having maximum conductivity of ~10⁻³ S cm⁻¹ at room temperature, which is acceptable from device fabrication point of view. Potential window and ionic transference number has been obtained to examine the potential limit and ionic characteristics of optimized GPE system. Temperature dependence behavior of electrical conductivity curve follows Arrhenius nature in the temperature range of 303–373 K. Pattern of dielectric constant and its loss as a function of frequency and temperature have been studied and is being explained on the basis of electrode interfacial polarization effect. Frequency-dependent conductivity spectra obey the Jonscher’s power law. Further, optimized composition of GPE has been tested successfully for its application in supercapacitor fabrication with activated charcoal as an electrode material. Maximum specific capacitance of 118.6 mF cm⁻² equivalent to single electrode specific capacitance of 61.7 F g⁻¹ have been observed for the optimized GPE film.

     

Development of Dissociation-Enhanced Lanthanide Fluoroimmunoassay for Measuring Leptin

Development of a dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) for measuring leptin, a satiety hormone of appetite control, was conducted in sandwich assay format exploiting a microplate immobilized with an anti-leptin antibody and another antibody raised against leptin and tagged with an europium chelate. In the leptin DELFIA of this study, amounts of antibody coated to the microplate and of the bioconjugate for the second immune reaction were optimized as 0.5 μg and 200 ng per well, respectively. When plotted in double-logarithmic scale, a linear relationship of y (log₁₀ response signal) = 0.6023× (log₁₀ leptin concentration) + 3.4084 (r² = 0.9646) was obtained at the leptin concentrations of 0.01─50 ng/mL with the limit of detection of 0.01 ng/mL. Individual leptin concentrations in various samples were well convergent to the calibration curve of the current assay. When applied to the measurement of leptin in a rat serum, the present assay was found quite effective and was competitive to a commercial sandwich-type ELISA.     

Observation of Brownian relaxation of magnetic nanoparticles using HTS SQUID

We analyzed the sensitivity of a separationless immunoassay scheme using functionalized magnetic nanoparticles (MNPs) and a sensitive HTS SQUID magnetometer. The signal of a 100 μL sample at a concentration of 1 mg/mL and field of 7.5 nT was 20 mΦ₀. This makes it possible for the sensitivity to be within the range of 50 ng/mL at the required time of up to 100 s per a point in the frequency spectrum.     

The use of the platinum electrode coated with ultrathin poly(allylamine hydrochloride)/Nafion films for selective detection of hydrogen peroxide

In this paper, a novel polyelectrolyte multilayer (PEM) film-coated platinum electrode for the selective detection of H₂O₂ was presented. The PEM film was formed by the layer-by-layer assembly technique. The quartz crystal microbalance experiments showed that the thickness of the prepared Nafion layer was about 8 nm and depended on the pH of poly(allylamine hydrochloride) solution. The combination of different polyanions and polycations layers was investigated, and it is found that ploy(allylamine hydrochloride) (PAH) and Nafion composited film functioned best as a diffusion barrier toward uric acid (UA) and ascorbic acid (AA) while allowed H₂O₂ to pass through smoothly. When the platinum electrode coated with two-bilayer film, (PAH/Nafion)₂, the amperometric responses of 0.1 mM UA and 0.1 mM AA were respectively 0.008 and 0.006 μA, which were only 0.2% or less of the response of 0.1 mM H₂O₂ (4.0 μA). The linear response range of the electrode toward H₂O₂ was from 1.0 μM to 1.0 mM, and the detection limit was 0.3 μM. The electrode also displayed high operational stability and long-term storage stability.     

2,6-Dansyl Azide as a Fluorescent Probe for Hydrogen Sulfide

A second-generation sulfonyl azide-based fluorescent probe, 2,6-DNS-Az, has been developed for the quantitative detection of H₂S in aqueous media such as phosphate buffer and bovine serum. Compare to the first-generation 1,5-DNS-Az probe, this probe shows both high sensitivity in phosphate buffer without the need for addition of surfactant and selectivity for sulfide over other anions and biomolecules, and thus can be used as a useful tool for detection of H₂S in the biological system.     

Production of anti-fullerene C<Subscript>60</Subscript> polyclonal antibodies and study of their interaction with a conjugated form of fullerene

The aim of this study was to produce anti-fullerene C₆₀ antibodies for the development of detection systems for fullerene C₆₀ derivatives. To produce anti-fullerene C₆₀ antibodies, conjugates of the fullerene C₆₀ carboxylic derivative with thyroglobulin, soybean trypsin inhibitor, and bovine serum albumin were synthesized by carbodiimide activation and characterized. Immunization of rabbits by the conjugates led to the production of polyclonal anti-fullerene antibodies. The specificity of the immune response to fullerene was investigated. Indirect competitive immunoenzyme assay was developed for the determination of conjugated fullerene with detection limits of 0.04 ng/mL (calculated for coupled C₆₀) and 0.4 ng/mL (accordingly to total fullerene–protein concentration).     

Electrochemical immune bioassay for the antigen–antibody interaction based on [Fe(CN)<Subscript>6</Subscript>]<Superscript>4-/3-</Superscript> and [AuCl<Subscript>4</Subscript>]<Superscript>-</Superscript> ions-derivated biomimetic interface

A simple and sensitive electrochemical immune bioassay for the detection of hepatitis B surface antigen (HBsAg), as a model, was developed based on [Fe(CN)₆]^4-/3- and [AuCl₄]^- ions-derivated biomimetic interface in this study. A layer of [Fe(CN)₆]^4-/3- film (i.e., Prussian blue, PB) was initially electrodeposited onto a glassy carbon electrode, and then [AuCl₄]^- ions were reduced under the potentiostat to form gold nanoparticles on the PB film. Finally, hepatitis B surface antibody (HBsAb) was adsorbed onto the nanogold surface. The performance and factors influencing the immunosensor were assessed and optimized. The proposed immunosensor exhibits a specific response to HBsAg in the range of 2.13–314.3 ng∙ml^-1 with a detection limit of 0.42 ng∙ml^-1. In addition, the developed immunosensor shows high sensitivity, good reproducibility, and long-term stability. Importantly, the ions-derivated biomimetic interface could be further extended for the immobilization of other proteins and biocompound.     

A new tyrosinase biosensor based on covalent immobilization of enzyme on N-(3-aminopropyl) pyrrole polymer film

A conducting polymer film of N-amino substituted pyrrole monomer has been prepared for covalent immobilization of enzyme for biosensing applications, illustrated by tyrosinase (PPO). The tyrosinase enzyme retains its bioactivity when being immobilized on N-substituted pyrrole polymer film by covalent bonding. The enzyme electrode was characterized by UV–Vis and infrared spectroscopy. Phenolic compounds were quantitatively estimated by the direct electrochemical reduction of enzymatically liberated quinone species at −0.2 V vs. Ag/AgCl. The results of amperometric response measurements conducted on enzyme electrode show sensitivity of 57.6, 71.4 and 45.8 mA M⁻¹ cm⁻² and a linear response range of 1.8–170.2, 1.3–110.1 and 2.1–168 μM for phenol, catechol and p-cresol, respectively. The biosensor exhibits a lowest detection limit of 0.9, 0.7 and 1.1 μM, for phenol, catechol and p-cresol, respectively and a period of stable sensitivity of 3 months at 4–5 °C.     

A sonochemical synthesis of SrTiO3 supported N-doped graphene oxide as a highly efficient electrocatalyst for electrochemical reduction of a chemotherapeutic drug

A sonochemical based green synthesis method playa powerful role in nanomaterials and composite development. In this work, we developed a perovskite type of strontium titanate via sonochemical process. SrTiO₃ particles were incorporated with nitrogen doped graphene oxide through simple ultrasonic irradiation method. The SrTiO₃/NGO was characterized by various analytical methods. The nanocomposite of SrTiO₃/NGO was modified with laser-induced graphene electrode (LIGE). The SrTiO₃/NGO/LIGE was applied for electrochemical sensor towards chemotherapeutic drug detection (nilutamide). Cyclic voltammetry (CV) and differential pulse voltammetry (DPV) techniques have been used to examine the electrochemical performance of nilutamide (anti-cancer drug). DPV was found to be more sensitive and found to exhibit a sensitivity 8.627 µA µM⁻¹ cm⁻² for SrTiO₃/NGO/LIGE with a wide linear range (0.02–892 µM) and low Limit of detection (LOD: 1.16 µM). SrTiO₃/NGO/LIGE has been examined for the detection of nilutamide in blood serum and urine samples and obtained a good recovery in the range of 97.2–99.72 %. The enhanced stability and selectivity and practical application results indicates the suitability of SrTiO₃/NGO/LIGE towards the detection of nilutamide drug in pharmaceutical industries.     

SILAR deposited Bi2S3 thin film towards electrochemical supercapacitor

Bi₂S₃ thin film electrode has been synthesized by simple and low cost successive ionic layer adsorption and reaction (SILAR) method on stainless steel (SS) substrate at room temperature. The formation of interconnected nanoparticles with nanoporous surface morphology has been achieved and which is favourable to the supercapacitor applications. Electrochemical supercapacitive performance of Bi₂S₃ thin film electrode has been performed through cyclic voltammetry, charge-discharge and stability studies in aqueous Na₂SO₄ electrolyte. The Bi₂S₃ thin film electrode exhibits the specific capacitance of 289 Fg⁻¹ at 5 mVs⁻¹ scan rate in 1 M Na₂SO₄ electrolyte.     

Surface-enhanced Raman scattering study of carcinoembryonic antigen in serum from patients with colorectal cancers

In this work, we developed a SERS platform for quantitative detection of carcinoembryonic antigen (CEA) in serum of patients with colorectal cancers. Anti-CEA-functionalized 4-mercaptobenzoic acid–labeled Au/Ag core-shell bimetallic nanoparticles were prepared first and then used to analyze CEA antigen solutions of different concentrations. A calibration curve was established in the range from 5 × 10^?3 to 5 × 10⁵ ng/mL. Finally, this new SERS probe was applied for quantitative detection of CEA in serum obtained from 26 colorectal cancer patients according to the calibration curve. The results were in good agreement with that obtained by electrochemical luminescence method, suggesting that SERS immunoassay has high sensitivity and specificity for CEA detection in serum. A detection limit of 5 pg/ml was achieved. This study demonstrated the feasibility and great potential for developing this new technology into a clinical tool for analysis of tumor markers in the blood.